池蝶蚌Sox2 基因在不同组织及不同月龄精巢中的表达

Sox 基因家族在胚胎发育过程和性别分化中起重要作用, 为研究池蝶蚌中Sox 基因的功能, 以人SRY基因HMG-box 保守区的序列设计简并引物, 以雌、雄池蝶蚌基因组DNA 和精巢cDNA 为模板进行扩增, 获得了2 个不完全相同的序列, 分别为DNA-HMG1、DNA-HMG2 和cDNA-HMG, 长度均为220 bp, 编码73个氨基酸。与人等物种Sox1、Sox2、Sox3 及Sox14 有很高的同源性, 雌雄个体之间没有序列差异性。采用RACE-PCR 扩增获得了池蝶蚌性腺Sox2 部分cDNA 片段, 长度为1774 bp, 该序列核苷酸与欧洲帽贝的SoxB和人类的Sox2 的同源性最高; 在部分开放阅读框249 个氨基酸残基中, 具有Sox 家族典型的HMG-box 结构域, 与人类、小鼠、原鸡和斑马鱼等Sox2 的HMG-box 同源性为98%。为了解该基因在各组织中的表达情况,采用实时荧光定量PCR 方法分析了外套膜、闭壳肌、鳃、肠、肝、肾、精巢和卵巢在内的8 种组织hs-Sox2的表达情况, 结果显示, hs-Sox2 基因在8 种组织中均有表达, 其中在肾脏中的表达量最高, 其次是肠与闭壳肌, 在雄性性腺中的表达量明显高于雌性性腺, 在肝脏中的表达量最低; 为了解hs-Sox2 在不同性腺发育时期的表达情况, 采用实时荧光定量PCR 方法分析了5 个不同月龄的精巢组织中hs-Sox2 的表达情况, 结果显示在39 月龄性腺的表达量最高, 其次是16 月龄性腺, 63 月龄蚌中的表达量最少。以上结果表明, hs-Sox2 基因可能参与了池蝶蚌精巢的发育及功能的维持。       

Cloning and functional analysis of the Sry-related HMG box gene, Sox18

The Sox gene family (Sry like HMG box gene) is characterised by a conserved DNA sequence encoding a domain of approximately 80 amino acids which is responsible for sequence specific DNA binding. We initially published the identification and partial cDNA sequence of murine Sox18, a new member of this gene family, isolated from a cardiac cDNA library. This sequence allowed us to classify Sox18 into the F sub-group of Sox proteins, along with Sox7 and Sox17. Recently, we demonstrated that mutations in the Sox18 activation domain underlie cardiovascular and hair follicle defects in the mouse mutation, ragged (Ra) (Pennisi et al., 2000. Mutations in Sox18 underlie cardiovascular and hair follicle defecs in ragged mice. Nat. Genet. 24, 434-437). Ra homozygotes lack vibrissae and coat hairs, have generalised oedema and an accumulation of chyle in the peritoneum. Here we have investigated the genomic sequences encoding Sox18. Screening of a mouse genomic phage library identified four overlapping clones, we sequenced a 3.25 kb XbaI fragment that defined the entire coding region and approximately 1.5 kb of 5' flanking sequences. This identified (i) an additional 91 amino acids upstream of the previously designated methionine start codon in the original cDNA, and (ii) an intron encoded within the HMG box/DNA binding domain in exactly the same position as that found in the Sox5, -13 and -17 genes. The Sox18 gene encodes a protein of 468 aa. We present evidence that suggests HAF-2, the human HMG-box activating factor -2 protein, is the orthologue of murine Sox18. HAF-2 has been implicated in the regulation of the Human IgH enhancer in a B cell context. Random mutagenesis coupled with GAL4 hybrid analysis in the activation domain between amino acids 252 and 346, of Sox18, implicated the phosphorylation motif, SARS, and the region between amino acid residues 313 and 346 as critical components of Sox18 mediated transactivation. Finally, we examined the expression of Sox18 in multiple adult mouse tissues using RT-PCR. Low-moderate expression was observed in spleen, stomach, kidney, intestine, skeletal muscle and heart. Very abundant expression was detected in lung tissue.     

Sox8基因HMG盒区内含子剪接位点分析

Sox基因参与广泛的发育调控过程.为了确定Sox8基因HMG盒区内含子的大小及剪接位点,通过计算机分析本室克隆得到的泥鳅Sox8基因包括HMG盒区在内的一段基因组序列,推测在HMG盒区可能存在一个内含子,进一步通过RT-PCR方法,克隆泥鳅Sox8基因HMG盒区cDNA片段,与基因组序列比较分析,确认在泥鳅Sox8基因的HMG盒区存在一个内含子,并确定了该内含子的序列及剪切位点.同时,比较分析了人和泥鳅Sox8在HMG盒区的内含子的剪切位点.结果显示,Sox8基因HMG盒区内含子剪切位点在进化上是保守的。 Abstract:Sox genes have diverse roles in developing processes,it was inferred there may be one intron in the HMG box by analyzing the genomic DNA sequence of the paramisgurnus dabryanus Sox8 gene.RT-PCR was used to verify the intron and its splicing site.First strand of the cDNA reverse-transcribed from liver RNA was amplified using the primers flanking the HMG box.RT-PCR products were cloned into the pUC18 plasmid and sequenced.The eomparison between the cDNA and genomic DNA sequence revealed the splicing site of the intron existing in the HMG-box of the paramisgurnus dabryanus Sox8 gene.We also compared the splicing site of the intron in the HMG-box of Sox8 gene between paramisgurnus dabryanus and human.These results suggest that the splicing site in the HMG-box of Sox8 gene was conserved.     

Group B sox genes that contribute to specification of the vertebrate brain are expressed in the apical organ and ciliary bands of hemichordate larvae

We have identified and characterized the sequence and expression of two Group B Sox genes in the acorn worm, Ptychodera flava. One sequence represents a Group B1 Sox gene and is designated Pf-SoxB1; the other is a Group B2 Sox gene and is designated Pf-SoxB2. Both genes encode polypeptides with an HMG domain in the N-terminal half. Whole-mount in situ hybridization to embryonic and larval stages of P. flava shows that the two genes are expressed in rather similar patterns at these stages. Expression is first detected in the cells of the blastula and subsequently localizes to the ectoderm during gastrulation. As the mouth forms, expression becomes concentrated in the stomodeum region. During morphogenesis of the tornaria larva, expression in the stomodeal ectoderm remains prominent around the mouth and under the oral hood. Later the genes are prominently upregulated in the ciliary bands and the apical organ. These results provide additional evidence that genes playing essential roles in the formation of the chordate dorsal central nervous system function in the development of the ciliary bands and apical organ, neural structures of this non-chordate deuterostome larva.     

Sox15 is up regulated in the embryonic mouse testis

The mammalian sex determining region on the Y chromosome, SRY, is the founding member of the SOX gene family. SOX genes share a common DNA-binding motif termed the HMG box and have diverse roles in vertebrate embryonic development and tissue differentiation. Sox15 expression was analysed during mouse embryogenesis by whole-mount in situ hybridisation and Real Time RT-PCR. Sox15 was found to be expressed in developing mouse gonads from 11.5 dpc to 13.5 dpc with a peak of expression at 12.5 dpc. Expression was approximately twice as high in the male gonad as in the female gonad.     

A Drosophila group E Sox gene is dynamically expressed in the embryonic alimentary canal

We have identified a novel Drosophila Sox-domain gene, Sox100B, related to the vertebrate group E genes Sox9 and Sox10. In vertebrates, group E Sox genes are expressed in the developing gonad, adult kidney and gut as well as other tissues. During embryogenesis in Drosophila, Sox100B is expressed in two rows of large intestinal cells, in midgut basophilic cells, in the Malpighian tubules and at the posterior cap of gonadal mesoderm. Our observations indicate that aspects of tissue-specific expression, as well as sequence, are conserved between vertebrate and invertebrate group E Sox proteins.